Download Free Carbon Nanotube Based Sensors For Label Free Protein Detection Book in PDF and EPUB Free Download. You can read online Carbon Nanotube Based Sensors For Label Free Protein Detection and write the review.

Optical biosensors based on fluorescent single-walled carbon nanotubes (SWNT) are a promising alternative to conventional biosensors due to the exceptional photophysical properties of SWNT. Such sensors can enable highly-sensitive, selective, and real-time detection of biological analytes. However, important questions regarding sensor fabrication and reproducibility must be addressed for these sensors to be of practical value. Herein we describe the use of highly-purified, single-chirality SWNT which are functionalized for antibody detection, and demonstrate that reproducibility is drastically improved with these SWNT. Further, we observe a concentration dependence of the effective equilibrium dissociation constant, KD,eff, which is in good agreement with previous reports, yet has eluded mechanistic description due to complexities associated with multivalent interactions. We show that a bivalent binding mechanism is able to describe this concentration dependence of KD,eff which varies from 100 pM to 1 uM for IgG concentrations from 1 ng/ml to 100 ug/ml, respectively. The mechanism is shown to describe the unusual concentration-dependent scaling demonstrated by other sensor platforms in the literature, and a comparison is made between resulting parameters. The platform is then extended to the detection of human growth hormone (hGH) using SWNT functionalized with a native hGH receptor (hGH-R), with potential use as a real-time and label-free measurement of protein activity. Native hGH is detected in the micromolar range, and an invariant equilibrium dissociation constant of 9 uM is revealed upon fitting the calibration curve to a single-site adsorption model. Selective detection of native hGH over thermally denatured hGH is shown at a concentration which is 1% of a clinical dose. Lastly, a multichannel detector was built to demonstrate real-time characterization of multiple protein properties. This work could find broad impact in biomanufacturing as real-time analysis of complex biologics is a long-standing goal in this field.
Nanoengineered glycan sensors may help realize the long-held goal of accurate and rapid glycoprotein profiling without labeling or glycan liberation steps. Current methods of profiling oligosaccharides displayed on protein surfaces, such as liquid chromatography, mass spectrometry, capillary electrophoresis, and microarray methods, are limited by sample pretreatment and quantitative accuracy. Microarrayed platforms can be improved with methods that better estimate kinetic parameters rather than simply reporting relative binding information. These quantitative glycan sensors are enabled by an emerging class of nanoengineered materials that differ in their mode of signal transduction from traditional methods. Platforms that respond to mass changes include a quartz crystal microbalance and cantilever sensors. Electronic response can be detected from electrochemical, field effect transistor, and pore impedance sensors. Optical methods include fluorescent frontal affinity chromatography, surface plasmon resonance methods, and fluorescent single walled carbon nanotubes-(SWNT). Advantages of carbon nanotube sensors include their sensitivity and ability to multiplex. The focus of this work has been to develop carbon nanotube-based sensors for glycans and proteins. Before detailing the development of these new sensors, the thesis will begin with a very brief primer on glycobiology, its connection to medicine, and the advantages and limitations of existing tools for glycan analysis. In the second chapter we model the use of quantitative nanosensors in a weak affinity dynamic microarray (WADM) to simulate practical uses of these sensors in bioprocessing and clinical diagnostics. There is significant interest in developing new detection platforms for characterizing glycosylated proteins, despite the lack of easily synthesized model glycans or high affinity receptors for this analytical problem. In the third chapter we experimentally demonstrate 'proof of concept' of carbon nanotubebased glycan sensors. This is done with a sensor array employing recombinant lectins as glycan recognition sites tethered via Histidine tags to Ni2l complexes that act as fluorescent quenchers for SWNT embedded in a chitosan hydrogel spot to measure binding kinetics of model glycans. We examine as model glycans both free and streptavidin-tethered biotinylated monosaccharides. Two higher-affined glycan-lectin pairs are explored: fucose (Fuc) to PA-IIL and N-acetylglucosamine (GlcNAc) to GafD. The dissociation constants (KD) for these pairs as free glycans (106 and 19 [mu]M respectively) and streptavidin-tethered (142 and 50 [mu]M respectively) were found. The absolute detection limit for the first-generation platform was found to be 2 pg of glycosylated protein or 100 ng of free glycan to 20 pg of lectin. Glycan detection (GlcNAc-streptavidin at 10 [mu]M) is demonstrated at the single nanotube level as well by monitoring the fluorescence from individual SWNT sensors tethered to GafD lectin. Over a population of 1000 nanotubes, 289 of the SWNT sensors had signals strong enough to yield kinetic information (KD of 250 ± 10 [mu]M). We are also able to identify the locations of "strong-transducers" on the basis of dissociation constant (4 sensors with KD 10 [Mu]) or overall signal modulation (8 sensors with 5% quench response). We report the key finding that the brightest SWNT are not the best transducers of glycan binding. SWNT ranging in intensity between 50 and 75% of the maximum show the greatest response. The ability to pinpoint strong-binding, single sensors is promising to build a nanoarray of glycan-lectin transducers as a high throughput method to profile glycans without protein labeling or glycan liberation pretreatment steps. In the fourth chapter we move from detection of model glycoproteins (streptavidin with biotinylated glycans) to a more applied problem: detection of antibodies and their glycosylation. We do this with a second generation array of SWNT nanosensors in an array format. It is widely recognized that an array of addressable sensors can be multiplexed for the label-free detection of a library of analytes. However, such arrays have useful properties that emerge from the ensemble, even when monofunctionalized. As examples, we show that an array of nanosensors can estimate the mean and variance of the observed dissociation constant (KD), using three different examples of binding IgG with Protein-A as the recognition site, including polyclonal human IgG (KD [mu] = 19 [mu]M, [sigma]2 = 1000 [mu]M2 ). murine IgG (KD = 4.3 [mu]M, 2= 3 [mu]M 2), and human IgG from CHO cells (KD [mu] = 2.5 nM, [sigma]F2 = 0.01 RM2). Second, we show that an array of nanosensors can uniquely monitor weakly-affined analyte interactions via the increased number of observed interactions. One application involves monitoring the metabolically-induced hypermannosylation of human IgG from CHO using PSA-lectin conjugated sensor arrays where temporal glycosylation patterns are measured and compared. Finally, the array of sensors can also spatially map the local production of an analyte from cellular biosynthesis. As an example we rank productivity of IgG-producing HEK colonies cultured directly on the array of nanosensors itself. One great limitation to these practical applications, common to other new sensor developments, are the constraints of large, bulky, and capital-intensive excitation sources, optics, and detectors. In the fifth chapter we detail the design of a lightweight, field-portable detection platform for SWNT based sensors using stock parts with a total cost below $3000. The portable detector is demonstrated with antibody detection in our lab and onsite at a commercial facility 3700 miles away with complex production samples. Along the course of developing these sensors, there was a need to analyze noisy data sets from signal nanotubes (Chapter 3) to determine distinct binding states. NoRSE was developed to analyze highfrequency data sets collected from multi-state, dynamic experiments, such as molecular adsorption and desorption onto carbon nanotubes. As technology improves sampling frequency, these stochastic data sets become increasingly large with faster dynamic events. More efficient algorithms are needed to accurately locate the unique states in each time trace. NoRSE adapts and optimizes a previously published noise reduction algorithm (Chung et al., 1991) and uses a custom peak flagging routine to rapidly identify unique event states. The algorithm is explained using experimental data from our lab and its fitting accuracy and efficiency are then shown with a generalized model of stochastic data sets. The algorithm is compared to another recently published state finding algorithm and is found to be 27 times faster and more accurate over 55% of the generalized experimental space. This work is detailed in Chapter 6. Future uses of these sensors include in vivo reporters of protein biomarkers. In Chapter 7, three-dimensional tracking of single walled carbon nanotubes (SWNT) with an orbital tracking microscope is demonstrated for this purpose. We determine the viscosity regime (above 250 cP) at which the rotational diffusion coefficient can be used for length estimation. We also demonstrate SWNT tracking within live HeLa cells and use these findings to spatially map corral volumes (0.27-1.32 Im 3), determine an active transport velocity (455 nm/s), and calculate local viscosities (54-179 cP) within the cell. With respect to the future use of SWNTs as sensors in living cells, we conclude that the sensor must change the fluorescence signal by at least 4-13% to allow separation of the sensor signal from fluctuations due to rotation of the SWNT when measuring with a time resolution of 32 ms. In the final chapter we draw conclusions from the development of this carbon nanotube-based sensor for glycan analysis and show the start of future work with arrays of SWNT sensors for glycoprofiling.
This volume summarizes the state-of-the-art technologies, key advances and future trends in the field of label-free biosensing. It provides detailed insights into the different types of solid-state, label-free biosensors, their underlying transducer principles, advanced materials utilized, device-fabrication techniques and various applications. The book offers graduate students, academic researchers, and industry professionals a comprehensive source of information on all facets of label-free biosensing and the future trends in this flourishing field. Highlights of the subjects covered include label-free biosensing with: · semiconductor field-effect devices such as nanomaterial-modified capacitive electrolyte-insulator-semiconductor structures, silicon nanowire transistors, III-nitride semiconductor devices and light-addressable potentiometric sensors · impedimetric biosensors using planar and 3D electrodes · nanocavity and solid-state nanopore devices · carbon nanotube and graphene/graphene oxide biosensors · electrochemical biosensors using molecularly imprinted polymers · biomimetic sensors based on acoustic signal transduction · enzyme logic systems and digital biosensors based on the biocomputing concept · heat-transfer as a novel transducer principle · ultrasensitive surface plasmon resonance biosensors · magnetic biosensors and magnetic imaging devices
As electronics reach nanometer size scales, new avenues of integrating biology and electronics become available. For example, nanoscale field-effect transistors have been integrated with single neurons to detect neural activity. Researchers have also used nanoscale materials to build electronic ears and noses. Another exciting development is the use of nanoscale biosensors for the point-of-care detection of disease biomarkers. This thesis addresses many issues that are relevant for electrical sensing applications in biological environments. As an experimental platform we have used carbon nanotube field-effect transistors for the detection of biological proteins. Using this experimental platform we have probed many of properties that control sensor function, such as surface potentials, the response of field effect transistors to absorbed material, and the mass transport of proteins. Field effect transistor biosensors are a topic of active research, and were first demonstrated in 1962. Despite decades of research, the mass transport of proteins onto a sensor surface has not been quantified experimentally, and theoretical modeling has not been reconciled with some notable experiments. Protein transport is an important issue because signals from low analyte concentrations can take hours to develop. Guided by mass transport modeling we modified our sensors to demonstrate a 2.5 fold improvement in sensor response time. It is easy to imagine a 25 fold improvement in sensor response time using more advanced existing fabrication techniques. This improvement would allow for the detection of low concentrations of analyte on the order of minutes instead of hours, and will open the door point-of-care biosensors.
Biosensors are poised to make a large impact in environmental, food, and biomedical applications, as they clearly offer advantages over standard analytical methods, including minimal sample preparation and handling, real-time detection, rapid detection of analytes, and the ability to be used by non-skilled personnel. Covering numerous applications of biosensors used in food and the environment, Portable Biosensing of Food Toxicants and Environmental Pollutants presents basic knowledge on biosensor technology at a postgraduate level and explores the latest advances in chemical sensor technology for researchers. By providing useful, state-of-the-art information on recent developments in biosensing devices, the book offers both newcomers and experts a roadmap to this technology. In the book, distinguished researchers from around the world show how portable and handheld nanosensors, such as dynamic DNA and protein arrays, enable rapid and accurate detection of environmental pollutants and pathogens. The book first introduces the basic principles of biosensing for newcomers to the technology. It then explains how the integration of a "receptor" can provide analytically useful information. It also describes trends in biosensing and examines how a small-sized device can have portability for the in situ determination of toxicants. The book concludes with several examples illustrating how to determine toxicants in food and environmental samples.
We have achieved two types of biomolecular sensors, colorimetric protein chips and label-free electrical sensors using high yield of single-walled carbon nanotubes (SWNTs). First, pseudo 3-dimensional SWNT films coated with carbonyldiimidazole-Tween20 (CDI-Tween20) surfactant demonstrated as a facile platform for fluorescence based protein chip. Highly selective protein bindings of biotin-BSA+SA (streptavidin) and SpA (protein A)+IgG (Immunoglobulin G) pairs, as well as those of small molecules such as FLAG peptide+anti-FLAG and biotin-SA. In this system, the geometry of pseudo 3-dimensional high yield of SWNTs preserves protein shapes intact, therefore increases the efficiencies of specific bindings. Furthermore, CDI-Tween20 mediates effective immobilization of probe proteins through covalent linkage, as well as prohibition of nonspecific bindings. By avoiding bovine serum albumin (BSA) which has been generally used as a biomolecular blocking agent during the protein chip manufacturing, it has been possible to reduce process steps, quenching of interaction signals from small molecules, and background noise. We also have fabricated ultrahigh sensitive electrical protein sensors using single-walled carbon nanotube-field effect transistors (SWNT-FETs) which contain increased Schottky contact area. A simple fabrication technique utilizing thin shadow mask and thermal evaporation at tilted angles allowed metal to penetrate underneath of the mask efficiently. Hence, thin and wide metal to SWNT contact regions are obtained, which could accommodate more proteins comparing to the typically fabricated SWNT-FET devices by photolithography. Direct protein adsorption of SpA and specific binding of hCG+anti Beta-hCG pairs on SWNT-FET changed more than 1% of conductance change at 1 pM concentration without NSB. These new SWNT-FET devices have increased the detection limit about four orders of magnitude compared to the previous devices.
Semiconducting single-walled carbon nanotubes (SWCNTs) are attractive transducers for biosensor applications due to their unique photostability, single molecule sensitivity, and ease of multiplexing. Sensors can be rendered selective via several detection modalities including the use of natural recognition elements (e.g., proteins) as well as the formation of synthetic molecular recognition sites from adsorbed heteropolymers. However, to date, deployment of SWCNT-based biosensors has been limited. The aim of this thesis was to study the design and development of SWCNT-based optical sensors for analytes relevant to the food and pharmaceutical industries including neurotransmitters, proteins, and metal ions. The research described in this thesis spans several levels of nanosensor development including: i) the fundamental study of SWCNT-polymer interactions and their dependence on solution properties; ii) sensor development using existing detection modalities and the use of mathematical modeling to guide sensor design and interpret data; and iii) the invention of a new sensor form factor enabling long-term sensor stability and point-of-use measurements. Our fundamental work on SWCNT-polymer interactions investigates the influence of polymer structure, SWCNT structure, and solution properties on molecular recognition, using single-stranded DNA as a model polymer system. We find that specific ssDNA sequences are able to form distinct corona phases across SWCNT chiralities, resulting in varying response characteristics to a panel of biomolecule probe analytes. In addition, we find that ssDNA-SWCNT fluorescence and wrapping structure is significantly influenced by the solution ionic strength, pH, and dissolved oxygen in a sequence-dependent manner. We are able to model this phenomenon and demonstrate the implications of solution conditions on molecular recognition, modulating the recognition of riboflavin. These results provide insight into the unique molecular interactions between DNA and the SWCNT surface, and have implications for molecular sensing, assembly, and nanoparticle separations. In addition to our experimental work, we used mathematical modeling to guide sensor design for biopharmaceutical characterization. A mathematical formulation for glycoprotein characterization was developed as well as a dynamic kinetic model to describe the data output by a label-free array of non-selective glycan sensors. We use the formulated model to guide microarray design by answering questions regarding the number and type of sensors needed to quantitatively characterize a glycoprotein mixture. As a second example, we report the design of a novel, diffusion-based assay for the characterization of protein aggregation. Specifically, we design hydrogel-encapsulated SWCNT sensors with a tunable hydrogel layer to influence the diffusion of immunoglobulin G protein species of variable size, and we develop a combined model that describes both the diffusion of analyte and analyte-sensor binding. By measuring the sensor response to a series of well-characterized protein standards that have undergone varying levels of UV stress, we demonstrate the ability to detect protein aggregates at a concentration as low as one percent on a molar basis. Finally, we report the development of a new form factor for optical nanosensor deployment involving the immobilization of SWCNT sensors onto paper substrates. We find that SWCNT optical sensors can be immobilized onto many different paper materials without influencing sensor performance. Moreover, we pattern hydrophobic barriers onto the paper substrates to create 1-dimensional sensor arrays, or barcodes, that are used for rapid, multiplexed characterization of several metal ions including Pb(II), Cd(II) and Hg(II). In addition to providing a new form factor for conducting point-of-use sensor measurements, these findings have the potential to significantly enhance the functionality of SWCNT-based optical sensors by interfacing them with existing paper diagnostic technologies including the manipulation of fluid flow, chemical reaction, and separation.
This book provides a comprehensive summary of the status of emerging sensor technologies and provides a framework for future advances in the field. Chemical sensors have gained in importance in the past decade for applications that include homeland security, medical and environmental monitoring and also food safety. A desirable goal is the ability to simultaneously analyze a wide variety of environmental and biological gases and liquids in the field and to be able to selectively detect a target analyte with high specificity and sensitivity. The goal is to realize real-time, portable and inexpensive chemical and biological sensors and to use these as monitors for handheld gas, environmental pollutant, exhaled breath, saliva, urine, or blood, with wireless capability.In the medical area, frequent screening can catch the early development of diseases, reduce the suffering of patients due to late diagnoses, and lower the medical cost. For example, a 96% survival rate has been predicted in breast cancer patients if the frequency of screening is every three months. This frequency cannot be achieved with current methods of mammography due to high cost to the patient and invasiveness (radiation). In the area of detection of medical biomarkers, many different methods, including enzyme-linked immunsorbent assay (ELISA), particle-based flow cytometric assays, electrochemical measurements based on impedance and capacitance, electrical measurement of microcantilever resonant frequency change, and conductance measurement of semiconductor nanostructures, gas chromatography (GC), ion chromatography, high density peptide arrays, laser scanning quantitiative analysis, chemiluminescence, selected ion flow tube (SIFT), nanomechanical cantilevers, bead-based suspension microarrays, magnetic biosensors and mass spectrometry (MS) have been employed. Depending on the sample condition, these methods may show variable results in terms of sensitivity for some applications and may not meet the requirements for a handheld biosensor.
This book draws together recent data on both cytoplasmic and flagellar dyneins and the proteins they interact with, to give the reader a clear picture of what is currently known about the structure and mechanics of these remarkable macro-molecular machines. Each chapter is written by active researchers, with a focus on currently used biophysical, biochemical, and cell biological methods. In addition to comprehensive coverage of structural information gained by electron microscopy, electron cryo-tomography, X-ray crystallography, and nuclear magnetic resonance, this book provides detailed descriptions of mechanistic experiments by single-molecule nanometry.
Nano-inspired Biosensors for Protein Assay with Clinical Applications introduces the latest developments in nano-inspired biosensing, helping readers understand both the fundamentals and frontiers in this rapidly advancing field. In recent decades, there has been increased interest in nano-inspired biosensors for clinical application. Proteins, e.g. antigen-antibody, tumor markers and enzymes are the most important target in disease diagnosis, and a variety of biosensing techniques and strategies have been developed for protein assay. This book brings together all the current literature on the most recent advances of protein analysis and new methodologies in designing new kinds of biosensors for clinical diagnostic use. Provides a single source of information on the latest developments in the field of biosensors for protein analysis and clinical diagnosis Focuses on biosensors fabricated with nanomaterials and nanotechnology Gives detailed methodologies for designing and fabricating nano-inspired biosensors